RNAi‐mediated silencing of insulin receptor substrate 1 (IRS‐1) enhances tamoxifen‐induced cell death in MCF‐7 breast cancer cells

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Abstract
Insulin receptor substrate 1 (IRS‐1) is a major downstream signaling protein for insulin and insulin‐like growth factor I (IGF‐I) receptors, conveying signals to PI‐3K/Akt and ERK1/2 pathways. In breast cancer, IRS‐1 overexpression has been associated with tumor development, hormone‐independence and antiestrogen‐resistance. In part, these effects are related to potentiation of IRS‐1/PI‐3K/Akt signaling. In estrogen sensitive breast cancer cell lines, tamoxifen treatment reduces IRS‐1 expression and function; consequently, inhibiting IRS‐1/PI‐3K signaling. We tested whether anti‐IRS1 siRNA could inhibit growth and survival of estrogen‐sensitive MCF‐7 breast cancer cells, when used alone or in combination with TAM. Our results indicated: (a) out of four tested anti‐IRS1 siRNAs, two siRNAs reduced IRS‐1 protein by approximately three‐fold in both growing and IGF‐I‐stimulated cells without affecting a closely related protein, IRS‐2; (b) these effects paralleled IRS1 mRNA downregulation by approximately three‐fold, measured by quantitative real time‐polymerase chain reaction; (c) action of anti‐IRS1 siRNAs induced the apoptotic response, observed by altered mitochondrial membrane potential coupled with downregulation of NF‐κB target Bcl‐xL and reduced cell viability; (d) anti‐IRS1 siRNA treatment enhanced the cytotoxic effects of TAM by ∼20%. In summary, anti‐IRS1 RNAi strategy could become a potent tool to induce breast cancer cell death, especially if combined with standard TAM therapy. J. Cell. Biochem. 98: 440–450, 2006.